| Objective:To provide scientific basis for the prevention and control of influenza A(H3N2)for our country,the molecular characteristics and epidemic trends of hemagglutinin(HA)and neuraminidase(NA)genes of human infection with seasonal influenza A(H3N2)virus were analyzed in our country from 2007 to 2017,and clinical significance of Th22 cells was preliminary expored in patients with H3N2 virus infection.Methods:(1)The gene sequences of HA and NA of influenza A(H3N2)virus strains isolated from China(Mainland of China,Hong Kong,Taiwan and macau.)during2007~2017 were obtained from the GISAID database.The molecular characteristics of HA/NA,and epidemic trends of influenza A(H3N2)virus were analyzed by the bioinformatics methods;(2)Flow cytometry analysis was performed to detect the frequency of Th22 cells in peripheral blood mononuclear cell(PBMC)from patients with influenza A(H3N2)virus infection including severe(15 cases)and mild(20 cases),and 20 healthy controls(HC);The expression levels of Th22 cells-associated cytokines(IL-22?IL-23?IL-6 and TNF-?)and transcription factors(AhR and ROR?t)mRNA were tested by real-time PCR method;Plasma concentrations of Th22 cells-associated cytokines(IL-22?IL-23?IL-6 and TNF-?)were measured by Liquid chip analysis platform.Results:(1)From 2007 to 2017,the number of influenza A(H3N2)virus strains of China were 1991 in total,there were 1173 strains in mainland China,479 strains in Hong Kong,324 strains in Taiwan and only 15 strains in Macao.At most,there were 110cases in Guangdong,followed by 104 cases in Beijing.In general,human infection with influenza A H3N2 virus subtype in China showed a local epidemic from 2007 to2017.The number of virus strains in 2007~2017 was gradually increased,except for the sudden expansion in 2009.(2)From 2007 to 2017,the HA amino acid sequence analysis showed that the virus strains belonging to the 3C.2a.1 clade had been increasing year by year since2010,and had become the most important epidemic dominant strain since 2016.The homologies of HA amino acids in 22 reference strains were 94.3%~99.9%.The homologies of clade 3C.3,clade 2,clade 3C.2,clade 3C.3a,clade 3C.2a,clade 3B,clade 3A,clade 1,and clade 3C.2a.1 were 97.5%~100.0%,96.7%~100.0%,95.4%~100.0%,95.3%~100.0%,95.1%~100.0%,94.9%~100.0%,94.3%~100.0%,94.0%~100.0%and 93.3%~100.0%,respectively.Both of clade 3C.3b and clade 3C.1were 94.6%~100.0%.(3)From 2007 to 2017,the amino acid sequences analysis of NA showed that the homologies of 22 reference strains were 95.3%~100.0%.The homologies of clade3C.1,clade 3C.2a,clade 3C.3a,clade 3A,clade 3B,clade 3C.2,clade 1 and clade 3C.2a.1were98.3%~100.0%,98.0%~100.0%,97.8%~100.0%,97.2%~100.0%,96.7%~100.0%,95.5%~100.0%,95.4%~100.0%and 95.2%~100.0%,respectively.Both of clade 3C.3b and clade 3C.3 were 96.3%~100.0%.(4)From 2007 to 2017,mutations in key sites of HA protein such as N121K and G/R142K were observed;268 strains with N121K mutations mainly came from 2017strains;358 strains with G/R142K mutations mainly originated from the strains between 2016 and 2017.Of the five antigen epitopes,the variants of A and B regions were the most frequently involved.The resistance site mutations with H275Y occurred in only 3 strains of NA protein in 2016;Other key amino acid sites including E119V,and R292K were not varianted,the NA gene was highly conserved during2007~2017.(5)The results showed that the proportion of IL-22~+CD4~+Th22 cells in CD4~+T cells was significantly higher than that in HC group,and the mild group was higher than the severe group;The expression levels of Th22 cells related cytokines(IL-22,IL-23,IL-6,TNF-?)and transcription factor(AhR)mRNA were significantly higher than those in the HC group,and only expression levels of IL-22 mRNA were significantly different between mild and severe groups;The plasma levels of Th22cells related cytokines(IL-22,IL-23,IL-6,TNF-?)were significantly higher than those in the HC group,and only the IL-22 concentrations were significantly different between the mild and severe groups.Conclusion:(1)The clades of influenza A(H3N2)virus are diverse,and thedifferent clades coexist.(2)The dominant epidemic strains were different among different years,especially since 2016,the clade 3C.2a.1 strains of influenza A(H3N2)virus were the most predominant strains in China.(3)The partial amino acid site of HA frequently mutated,but the NA sequence was relatively highly conserved,and only a few strains showed resistance locus mutations;(4)The frequency of Th22 cells,and the expression levels of IL-22 and other related cytokines and transcription factors were significantly increased in patients with H3N2 infection,Th22 cell frequency and expressions of IL-22factor were significant differences between the mild and severe groups. |
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